A role for YAP-mediated regulation of invadopodia in HNSCC cells

Abstract

ABSTRACTThe objective of this study was to determine whether nuclear translocation of the transcriptional coactivator Yes-associated protein (YAP) was sensitive to extracellular matrix (ECM) rigidity and promoted Rho-associated kinase 2 (ROCK2) expression to affect invadopodia maturation and ECM degradation. ECM rigidity mimicking head and neck squamous cell carcinoma (HNSCC) tumor mechanical properties was simulated in vitro using a well-established model based on fibronectin-conjugated polyacrylamide gels (PAAs). The ratio of nuclear to cytoplasmic YAP and overall cellular ROCK2 levels were evaluated in HNSCC cells using quantitative immunofluorescence. YAP-mediated ROCK2 expression in HNSCC cells was determined using nested PCR and Western blot in response to the YAP inhibitor verteporfin. Invadopodia and ECM degradation were evaluated in HNSCC cells with siRNA-mediated inhibition of YAP using quantitative immunofluorescence in invadopodia assays. Both YAP nuclear translocation and ROCK2 cellular levels increased with ECM rigidity. Inhibition of YAP activity with verteporfin decreased ROCK2 gene and protein expression. Knockdown of YAP with siRNA inhibited the formation of mature invadopodia and ECM degradation but not total invadopodia (i.e., mature and immature or not degrading). Our study suggests that tumor-associated ECM rigidity can promote mechanically-induced transcriptional regulation to control proteolytic activity by affecting invadopodia maturation.